Patients with neurological diseases, including Parkinson's disease, Alzheimer's disease, brain tumors and most neurogenetic disorders, suffer from severe debilitating symptoms and lack of therapeutic options that provide curative treatment. The accumulated knowledge of specific genetic targets that can alter or reverse the natural history of central nervous system (CNS) diseases has rendered gene therapy an attractive therapeutic strategy [O'Mahony, A. M., et al., J Pharm Sci, 2013. 102(10): 3469-3484; Lentz, et al., Neurobiol Dis, 2012. 48(2): 179-188.]. Multiple preclinical and clinical studies have aimed to improve the delivery of nucleic acids to the CNS using leading viral or non-viral gene vectors with specific focus to enhancing the level and distribution of transgene expression throughout the brain tissue [O'Mahony, et al., J Pharm Sci, 2013. 102(10): 3469-3484; Perez-Martinez, et al., J Alzheimers Dis, 2012. 31(4): 697-710].
Viral gene vectors, though relatively efficient, have been limited by one or more drawbacks, including low packaging capacity, technical difficulties in scale-up, high cost of production [Thomas, et al., Nat Rev Genet, 2003. 4(5): 346-358.] and risk of mutagenesis [Olsen and Stein, N Engl J Med, 2004. 350(21): 2167-2179.]. Furthermore, despite the immune privileged nature of the CNS, neutralizing immune responses may occur secondary to repeated administrations or prior exposures [Lentz, et al., Neurobiol Dis, 2012. 48(2): 179-188; Xiao, X., et al., J Virol, 1996. 70(11): 8098-8108; Chirmule, N., et al., J Virol, 2000. 74(5): 2420-2425; Lowenstein, P. R., et al., Curr Gene Ther, 2007. 7(5): p. 347-60; Lowenstein, P. R., et al., Neurotherapeutics, 2007. 4(4): 715-724; Voges, J., et al., Ann Neurol, 2003. 54(4): 479-487.].
Non-viral gene vectors can offer an attractive alternate strategy for gene delivery without many of these limitations [O'Mahony, A. M., et al., J Pharm Sci, 2013. 102(10): 3469-3484]. Cationic polymer-based gene vectors provide a tailorable platform for DNA condensation and efficient gene transfer in vitro and in vivo. Their positive charge density allows for stable compaction of negatively charged nucleic acids [Sun, X. and N. Zhang, Mini Rev Med Chem, 2010, 10(2): 108-125; Dunlap, D. D., et al., Nucleic Acids Res, 1997. 25(15): 3095-3101] and protects them from enzymatic degradation [Kukowska-Latallo, J. F., et al., Hum Gene Ther, 2000. 11(10): 1385-1395.]. Also, the number of protonable amines provides increased buffering capacity that facilitates endosome escape via the “proton sponge effect”, leading to efficient transfection [Akinc, A., et al., J Gene Med, 2005. 7(5): 657-663]. A wide variety of cationic polymers have been developed for this purpose, offering gene vectors with diverse physicochemical profiles and in vivo behaviors [Mintzer, M. A. and E. E. Simanek. Chem Rev, 2009. 109(2): 259-302; Pathak, et al., Biotechnol J, 2009. 4(11): 1559-72.].
However, non-viral gene vectors still face a number of barriers prior to reaching the target cells in the brain [O'Mahony, et al., J Pharm Sci, 2013. 102(10): 3469-3484]. Various strategies have been developed to manipulate or bypass the blood brain barrier (BBB) [Jain, Nanomedicine (Lond), 2012. 7(8): 1225-33; Wohlfart, et al., J Control Release, 2012. 161(2): 264-273.], which is the primary barrier to the systemic delivery of gene vectors to the brain. These approaches include, but are not limited to, direct, local administration to the CNS [Patel, et al., Advanced Drug Delivery Reviews, 2012. 64(7):701-705] and reversible disruption of the BBB via focused ultrasound [Vykhodtseva, et al., Ultrasonics, 2008. 48(4): 279-296] or chemical reagents [Kroll, et al., Neurosurgery, 1998. 42(5): 1083-1099; discussion 1099-100.]. However, once beyond the BBB, the anisotropic and electrostatically charged extracellular matrix (ECM) found between brain cells has been widely recognized as another critical barrier [Nance, et al., Sci Transl Med, 2012. 4(149): 149ra119; Sykova, et al., Physiol Rev, 2008. 88(4): 1277-1340; Zamecnik, J., Acta Neuropathol, 2005. 110(5):435-442]. This ‘brain tissue barrier’, regardless of administration method, hampers widespread distribution of macromolecules and nanoparticles in the brain, thereby limiting their coverage throughout the disseminated target area of neurological diseases [Voges, J., et al., Ann Neurol, 2003. 54(4): 479-487; Nance, E. A., et al., Sci Transl Med, 2012. 4(149): 149ra119; Sykova, et al., Physiol Rev, 2008. 88(4): 1277-340; MacKay, et al., Brain Res, 2005. 1035(2): 139-153]. The ECM is rich in hyaluronan, chondroitin sulfate, proteoglycans, link proteins and tenascins and may provide a negatively charged adhesive barrier to the penetration of cationic polymeric gene vectors [Sykova, et al., Physiol Rev, 2008. 88(4): 1277-1340; Zimmermann, et al., Histochem Cell Biol, 2008. 130(4): 635-653]. Moreover, the pore size of the ECM imposes a steric barrier for the movement of nanoparticles in the CNS with non-adhesive 114 nm, but not 200 nm, particles able to penetrate the brain tissue [Nance, E. A., et al., Sci Transl Med, 2012. 4(149): p. 149ra119; Kenny, G. D., et al., Biomaterials, 2013. 34(36): 9190-9200. It has been shown that sub-100 nm nanoparticles exceptionally well-coated with hydrophilic and neutrally charged polyethylene glycol (PEG) rapidly diffuse in the brain ECM allowing the Widespread distribution of therapeutics [Nance, E. A., et al., Sci Transl Med, 2012. 4(149): p. 149ra119].
Convection enhanced delivery (CED) can be applied to further enhance the distribution of therapeutics by providing a pressure gradient during intracranial administration [Allard, et al., Biomaterials, 2009. 30(12): 2302-2318.]. However, CED is unlikely to provide a significant benefit if particles remain entrapped in the brain parenchyma due to adhesive interactions and/or steric obstruction. Thus, physicochemical properties of particles that allow unhindered diffusion in the brain parenchyma remain critical for achieving enhanced particle penetration following CED [Allard, et al., Biomaterials, 2009. 30(12): 2302-18; Kenny, et al., Biomaterials, 2013. 34(36): 9190-9200]. However, even following CED, the interactions between positively charged gene vectors and the negatively charged ECM, confine cationic nanoparticles to the point of injection and perivascular spaces, and limit their penetration into the brain parenchyma [MacKay, et al., Brain Res, 2005. 1035(2): 139-153; Kenny, et al., Biomaterials, 2013. 34(36): 9190-9200; Writer, et al., J Control Release, 2012. 162(2): p. 340-8.].
It is therefore an object of the present invention to provide optimized physicochemical properties of stable gene vectors that can penetrate the brain parenchyma thus improving distribution and transgene expression throughout the brain tissue.
It is a further object of the present invention to provide gene delivery vectors with a favorable safety profile.
It is a yet further object of the present invention to combine nanoparticles with delivery strategies that can further enhance their distribution and transgene expression in the tissue, especially the brain.